Otto Warburg published a paper in 1927 that described the accumulation of lactate in the veins of experimental rats that had growing cancer cells. He explained this observation by concluding that cancer cells obtained metabolic energy from an aerobic glycolysis (i.e. fermentation) mechanism. This was a new phenomenon that was unique to cancer because glycolysis was normally observed to occur under anaerobic conditions, which is known as the Pasteur Effect (where oxygen inhibits glycolysis). Interest in further investigation of the Warburg effect has increased significantly over the last ten years. Recent studies have uncovered some of the biochemical mechanisms associated with aerobic glycolysis and cancer.
Warburg devoted his life to measuring and understanding energy flow through biological systems. Some of his measurement techniques came from the same laboratory where precision blackbody radiation measurements were performed at the end of the 19th century. Max Planck used data from this lab in 1900 to derive his law of radiation and calculate the value of ‘quantum action’, h=6.63×10−34 J s, work that launched the development of quantum mechanics (the field of physics that is used to design and fabricate the laser devices discussed below). In addition to optical measurement techniques, Warburg developed and used manometers to measure oxygen consumption, and thus energy input, of a variety of biological systems. His experiments had the unifying theme of understanding chemical energy flow through a biological system. Much of our textbook understanding of biological metabolism can trace its origins to work in his laboratory.
Injured Respiration
In a paper titled “On the Origin of Cancer Cells” published in Science in 1956, Warburg offers this summary:
The irreversible injuring of respiration is followed, as the second phase of cancer formation, by a long struggle for existence by the injured cells to maintain their structure, in which part of the cells perish from lack of energy, while another part succeed in replacing the irretrievably lost respiration energy by fermentation energy. Because of the morphological inferiority of fermentation energy, the highly differentiated body cells are converted by this into undifferentiated cells that grow wildly—the cancer cells.
FIG. 1 illustrates the respiration process for a healthy cell. It begins with the breakdown of glucose to pyruvate through the glycolysis mechanism where a small amount of energy for cell function, as represented by two adenosine triphosphate (A TP) molecules generated for each glucose molecule, is produced. Pyruvate then enters the respiration mechanism by interacting with pyruvate dehydrogenase (POH) resulting in conversion of pyruvate to acetyl-CoA, which is the molecule that enters the tricarboxylic acid (TCA) (or Krebs) cycle. This cycle is very efficient, producing up to 36 ATP molecules for each glucose molecule.
FIG. 2 illustrates a cell with injured respiration. In this case pyruvate does not enter the Krebs cycle. Instead it is shown to form lactate, which is the molecule that Warburg measured to identify the fermentation phenotype for cancer cells. He also observed that the rate of glycolysis for this phenotype was much higher than when respiration is not impaired. As a result, growing cancer cells can still receive sufficient energy even though the ATP-producing mechanism is not as efficient. This higher rate of glycolysis provides the scientific basis for positron emission tomography (PET) scans that reveal images of cancerous tumors. The technique involves administering a radiolabeled glucose analog [18F]-fluorodeoxyglucose (FOG) where the high rate of glycolysis within tumors concentrates the positron-emitting fluorine isotope. The widespread clinical success of PET scans offers convincing experimental confirmation of the selective high uptake of glucose in invasive tumors.
Underlying Cause of Injured Respiration
Hans Krebs, who worked in Otto Warburg's laboratory as a postdoctoral associate from 1926 to 1930, uncovered a variety of cyclic mechanisms in biological systems including the Krebs cycle (also known by other names such as the TCA or citric acid cycle or oxidative phosphorylation) while at universities in Freiburg, Germany and Oxford, England. In his 1981 biography of Otto Warburg, Krebs offered this opinion on Warburg's cancer theory:
Warburg's ‘primary cause of cancer’—the replacement of respiration by fermentation—may be a symptom of the primary cause, but is not the primary cause itself. The primary cause is to be expected at the level of the control of gene expression, the minutiae of which are unknown though some of the principles involved are understood.
As mentioned above, pyruvate enters the respiration mechanism by interacting with PDH resulting in conversion of pyruvate to acetyl-CoA. Recent research has provided some minutiae, as suggested by Krebs 25 years earlier, of the underlying genetic cause of injured respiration. By performing a series of measurements with mouse embryo fibroblasts, it has been shown that hypoxia-inducible factor 1 (HIF-1), a genetic transcription factor that responds to decreases in oxygen supply, induces pyruvate dehydrogenase kinase 1 (PDK1). PDK1 inhibits PDH thus blocking pyruvate entry into the respiratory Krebs cycle. Pyruvate therefore remains in the cytoplasm where it forms lactate by NADH reduction—the Warburg Effect.
The underlying cause of injured respiration is therefore connected to HIF-1 transactivation of the gene encoding PDK1, which can be considered a gate-keeping enzyme that regulates the flow of pyruvate, a product of glycolysis, into the mitochondria for oxidation. If PDK1 is encoded then pyruvate builds up in the cytoplasm, and the cell is forced to rely on metabolic energy from glycolysis even if oxygen is available.
The successful measurement of gas phase acetaldehyde using a IV-VI semiconductor diode laser, was performed by Kamat et al wherein the laser operated in cw mode with a heat sink cooled to 101 K by a closed-cycle compressor. FIG. 5 shows the spectral region covered by the tuning of this laser, which was near the P-branch of the carbonyl stretch mode of acetaldehyde. FIG. 6 shows an acetaldehyde absorption feature between 1727.05 cm−1 and 1727.15 cm−1 measured with this laser. The acetaldehyde absorption feature highlighted in the shaded region of FIG. 6 consists of coupled vibrational and rotational modes for the molecule. As shown in FIG. 9, using a 100 meter long optical path length Herriott gas cell a minimum detection limit of 50 ppb with a 10 second sample integration time was demonstrated. The mid-IR instrument described in Kamat et al. operated at cryogenic temperatures, which required a bulky and costly closed-cycle compressor. Smith et al. described the measurement of acetaldehyde using a variation of the mass spectroscopy method, selected ion flow tube (SIFT) MS that removes acetaldehyde's mass interference with carbon dioxide. FIG. 8 from Smith shows measured acetaldehyde concentrations in the headspace of cultured lung cancer cells. In addition, they observed a decrease in pH and a corresponding increase in lactic acid in the cell culture medium. These results were consistent with the Warburg Effect where blockage of pyruvate from entering the respiratory Krebs cycle results in the generation of lactic acid and gas phase acetaldehyde.
Unfortunately there does not exist a sensitive enough, easily portable and cost effective detection mechanism for air-based analytes. The prior art techniques are either not sensitive enough or expensive and bulky, mostly due to the need to provide large bulky compressor or cooling systems. For example, the spectrometer used by Kamat required a closed cycle refrigeration system to cool the system to 101K.
There is a need for a new sensor technology that includes a mid-IR detection device with an improved cooling system which allows for the rapid and cost-effective detection of analytes in gaseous samples, for example at sensitivities that allow for enhanced detection and/or treatments of cancer.